Assessment of antibiotic residues and microbiological safety in Lebanese dairy products: Akkawi cheese, yogurt, and labneh, collected across Lebanese dairy industries

Antibiotic residues in dairy products and the resulting antibiotic-resistant bacteria represent a major global public health concern. This study evaluated the microbiological safety of 400 dairy samples collected over two consecutive years (2022 and 2023) from 16 dairy residue industries across four governorates in Lebanon. In addition, antibiotic residues were screened from 126 dairy product samples randomly chosen out of the 400. An optimized multi-class method for the identification and quantification of 30 antibiotics was developed using liquid chromatography-mass spectrometry. Results revealed that 48% of the analyzed samples were contaminated with at least one antibiotic residue, among which 33% showed co-occurrence of multiple antibiotic classes. Meanwhile, microbiological assessment of 400 dairy samples collected over 2 years from the same dairies revealed higher levels of contamination with total coliforms, Escherichia coli, and Staphylococcus aureus in Akkawi cheese samples collected during the first year (44%, 32%, and 15%, respectively) compared to those collected the second year (38.8%, 31%, and 0%). Moderate contamination levels were observed in Labneh (strained yogurt) samples collected during both seasons, with yeast (12%; 11%), molds (1.2%; 11%), total coliforms (6%; 11%), and Escherichia coli (3.7%; 11%). On the other hand, comparable levels of non-conformity were observed in yogurt samples across the two years, with contamination by yeast (7.5%; 6.2%), molds (3.75%; 6.2%), total coliforms (26%; 31.2%), E. coli (20%; 21%), and Staphylococcus aureus (1.25%; 0%). Escherichia coli isolates exhibited resistance to tetracycline while being 100% sensitive to ciprofloxacin and nalidixic acid. As for S. aureus isolates, they presented a high sensitivity to tetracyclines, doxycycline, and ciprofloxacin. These findings highlight the importance of implementing and maintaining good hygienic practices within dairy industries. Antimicrobial residues of oxytetracycline, tetracycline, ciprofloxacin, sulfamethazine, and doxycycline were detected at levels below the maximum residue limits.

Adeyemo AA, Gbadegesin RA, Onyemenem TN, Ekweozor CC, 1994. Urinary tract pathogens and antimicrobial sensitivity patterns in children in Ibadan, Nigeria. Ann Trop Paediatr 14:271-4. DOI: https://doi.org/10.1080/02724936.1994.11747728

Al-Kadamany E, Toufeili I, Khattar M, Abou-Jawdeh Y, Harakeh S, Haddad T, 2002. Determination of shelf life of concentrated yogurt (labneh) produced by in-bag straining of set yogurt using hazard analysis. J Dairy Sci 85:1023-30. DOI: https://doi.org/10.3168/jds.S0022-0302(02)74162-3

Alomirah H, Al‐Mazeedi H, Al‐Zenki S, Al‐Aati T, Al‐Otaibi J, Al‐Batel M, Sidhu J, 2007. Prevalence of antimicrobial residues in milk and dairy products in the state of Kuwait. J Food Qual 30:745-63. DOI: https://doi.org/10.1111/j.1745-4557.2007.00157.x

Amer AMM, Constable PD, Goudah A, El Badawy SA, 2012. Pharmacokinetics of tulathromycin in lactating goats. Small Rumin Res 108:137-43. DOI: https://doi.org/10.1016/j.smallrumres.2012.07.003

Anastassiades M, Lehotay SJ, Štajnbaher D, Schenck FJ, 2003. Fast and easy multiresidue method employing acetonitrile extraction/partitioning and “dispersive solid-phase extraction” for the determination of pesticide residues in produce. J AOAC Int 86:412-31. DOI: https://doi.org/10.1093/jaoac/86.2.412

Bergwerff AA, Schloesser J, 2003. Antibiotics and drugs: residue determination. In: Encyclopaedia of food science, food technology and nutrition. Academic Press, London, UK. DOI: https://doi.org/10.1016/B0-12-227055-X/00052-3

Cabizza R, Rubattu N, Salis S, Pes M, Comunian R, Paba A, Addis M, Testa MC, Urgeghe PP, 2017. Transfer of oxytetracycline from ovine spiked milk to whey and cheese. Int Dairy J 70:12-7. DOI: https://doi.org/10.1016/j.idairyj.2016.12.002

Carrique-Mas JJ, Hökeberg I, Andersson Y, Arneborn M, Tham W, Danielsson-Tham ML, Osterman B, Leffler M, Steen M, Eriksson E, Hedin G, Giesecke J, 2003. Febrile gastroenteritis after eating on-farm manufactured fresh cheese – an outbreak of listeriosis? Epidemiol Infect 130:79-86. DOI: https://doi.org/10.1017/S0950268802008014

CLSI, 2012. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; Approved standard—9th edition (No. CLSI document M07–A9), 9th ed. Clinical and Laboratory Standards Institute, Wayne, PA.

CLSI, 2013. Performance standards for antimicrobial susceptibility testing; Twenty-third informational supplement (No. CLSI document M100-S23), 23rd ed. Clinical and Laboratory Standards Institute, Wayne, PA.

Dib, H., Hajj Semaan, E., & Noureddine, Z. (2008). Caractéristiques chimiques et microbiologiques des fromages libanais issus d’industries locales. Lebanese Science Journal, 9(2), 37–46

El Kojok H, Khalil M, Hage R, Jammoul R, Jammoul A, El Darra N, 2022. Microbiological and chemical evaluation of dairy products commercialized in the Lebanese market. Vet World 15:2575-86. DOI: https://doi.org/10.14202/vetworld.2022.2575-2586

EL-Ziney MG, Al-Turki AI, 2007. Microbiological quality and safety assessment of camel milk (Camelus dromedaries) in Saudi Arabia (Qassim region). Appl Ecol Env Res 5:115-22. DOI: https://doi.org/10.15666/aeer/0502_115122

European Commission, 2010. Commission Regulation (EU) No 37/2010 of 22 December 2009 on pharmacologically active substances and their classification regarding maximum residue limits in foodstuffs of animal origin (No. Regulation (EU) No 37/2010). In: Official Journal, L 15, 20/01/2010.

European Food Safety Authority, European Centre for Disease Prevention and Control, 2018. The European Union summary report on antimicrobial resistance in zoonotic and indicator bacteria from humans, animals and food in 2016. EFSA J 16:e05182. DOI: https://doi.org/10.2903/j.efsa.2018.5182

European Parliament, Council of the European Union, 2001. Directive 2001/82/EC of the European Parliament and of the Council of 6 November 2001 on the Community code relating to veterinary medicinal products. In: Official Journal, L 311.

European Parliament, Council of the European Union, 2002. Regulation (EC) No 178/2002 of the European Parliament and of the Council of 28 January 2002 laying down the general principles and requirements of food law, establishing the European Food Safety Authority and laying down procedures in matters of food safety. In: Official Journal, L 031, 1/02/2002.

FDA, 2001. Bacteriological analytical manual: chapter 18 – yeasts, molds and mycotoxins. Center for Food Safety and Applied Nutrition, U.S. Department of Health and Human Services, Washington DC, USA.

Ferrini AM, Trenta S, Mannoni V, Rosati R, Coni E, 2010. Depletion of long-acting ampicillin in goat milk following intramuscular administration. J Agric Food Chem 58:12199-2203. DOI: https://doi.org/10.1021/jf102351p

Gaudin V, Hedou C, Rault A, Verdon E, 2010. Validation of a Five Plate Test, the STAR protocol, for the screening of antibiotic residues in muscle from different animal species according to European Decision 2002/657/EC. Food Addit Contam Part A Chem Anal Control Expo Risk Assess 27:935-52. DOI: https://doi.org/10.1080/19440041003677483

Gaurav A, Gill JPS, Aulakh RS, Bedi JS, 2014. ELISA based monitoring and analysis of tetracycline residues in cattle milk in various districts of Punjab. Vet World 7:26-9. DOI: https://doi.org/10.14202/vetworld.2014.26-29

Graham F, Paradis L, Bégin P, Paradis J, Babin Y, Des Roches A, 2014. Risk of allergic reaction and sensitization to antibiotics in foods. Ann Allergy Asthma Immunol 113, 329-30. DOI: https://doi.org/10.1016/j.anai.2014.06.029

Gündoğan N, Citak S, Turan E, 2006. Slime production, DNase activity and antibiotic resistance of Staphylococcus aureus isolated from raw milk, pasteurised milk and ice cream samples. Food Control 17:389-92. DOI: https://doi.org/10.1016/j.foodcont.2005.01.006

Hatakka M, Björkroth KJ, Asplund K, Mäki-Petäys N, Korkeala HJ, 2000. Genotypes and enterotoxicity of Staphylococcus aureus isolated from the hands and nasal cavities of flight-catering employees. J Food Prot 63:1487-91. DOI: https://doi.org/10.4315/0362-028X-63.11.1487

IFAD, 2017. Harmonized actions for livestock enhanced production and processing project (HALEPP) – detailed design report. IFAD, Rome, Italy.

ISO, 1999. Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration of coagulase-positive staphylococci (Staphylococcus aureus and other species) — Part 1: Technique using Baird-Parker agar medium. ISO Norm 6888-1:1999). International Standardization Organization ed., Geneva, Switzerland.

ISO, 2001. Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration of β-glucuronidase-positive Escherichia coli — Part 2: Colony-count technique at 44°C using 5-bromo-4-chloro-3-indolyl β-D-glucuronide. ISO Norm 16649-2:2001. International Standardization Organization ed., Geneva, Switzerland.

ISO, 2006. Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration of coliforms — Colony-count technique. ISO Norm 4832:2006. International Standardization Organization ed., Geneva, Switzerland.

ISO, 2010. Microbiology of food and animal feeding stuffs — Preparation of test samples, initial suspension and decimal dilutions for microbiological examination — Part 5: Specific rules for the preparation of milk and milk products. ISO Norm 6887-5:2010. International Standardization Organization ed., Geneva, Switzerland.

ISO, 2017. Microbiology of the food chain — Horizontal method for the detection and enumeration of Listeria monocytogenes — Part 1: Detection method. ISO Norm ISO 11290-1:2017. International Standardization Organization ed., Geneva, Switzerland.

Joubrane K, Jammoul A, Daher R, Ayoub S, El Jed M, Hneino M, El Hawari K, Al Iskandarani M, Daher Z, 2022. Microbiological contamination, antimicrobial residues, and antimicrobial resistance in raw bovine milk in Lebanon. Int Dairy J 134:105455. DOI: https://doi.org/10.1016/j.idairyj.2022.105455

Kabrite S, Bou-Mitri C, El Hayek Fares J, Hassan HF, Matar Boumosleh J, 2019. Identification and dietary exposure assessment of tetracycline and penicillin residues in fluid milk, yogurt, and labneh: a cross-sectional study in Lebanon. Vet World 12:527-34. DOI: https://doi.org/10.14202/vetworld.2019.527-534

Khan AZ, Khan A, Hayat CS, Munir Z, Ayaz U, 2004. Prevalence of mastitis in buffaloes and antibiotics sensitivity profiles of isolates. Pak J Life Soc Sci 2:73-5.

Lanz R, Kuhnert P, Boerlin P, 2003. Antimicrobial resistance and resistance gene determinants in clinical Escherichia coli from different animal species in Switzerland. Vet Microbiol 91:73-84. DOI: https://doi.org/10.1016/S0378-1135(02)00263-8

LIBNOR, 2012. Milk and milk products — microbiological limits (National Standard No. NL 510). Lebanese Standards Institution, Beirut, Lebanon.

Le Loir Y, Baron F, Gautier M, 2003. Staphylococcus aureus and food poisoning. Genet Mol Res 2:63-76.

Mayssoun Z, Nadine N, 2010. Influence of production processes in quality of fermented milk "Laban" in Lebanon. Health 2:381-9. DOI: https://doi.org/10.4236/health.2010.24057

Ministry of Agriculture, 2010. Regulating the sale of veterinary medicines, memo No. 112/1.

Oliver SP, Jayarao BM, Almeida RA, 2005. Foodborne pathogens in milk and the dairy farm environment: food safety and public health implications. Foodborne Pathog Dis 2:115-29. DOI: https://doi.org/10.1089/fpd.2005.2.115

Olsen SJ, Ying M, Davis MF, Deasy M, Holland B, Iampietro L, Baysinger CM, Sassano F, Polk LD, Gormley B, Hung MJ, Pilot K, Orsini M, Van Duyne S, Rankin S, Genese C, Bresnitz EA, Smucker J, Moll M, Sobel J, 2004. Multidrug-resistant Salmonella Typhimurium infection from milk contaminated after pasteurization. Emerg Infect Dis 10:932-5. DOI: https://doi.org/10.3201/eid1005.030484

Pogurschi E, Ciric A, Zugrav C, Patrascu D, 2015. Identification of antibiotic residues in raw milk samples coming from the metropolitan area of Bucharest. Agric Agric Sci Procedia 6:242-5. DOI: https://doi.org/10.1016/j.aaspro.2015.08.066

Quintanilla P, Beltrán MC, Peris B, Rodríguez M, Molina MP, 2018. Antibiotic residues in milk and cheeses after the off-label use of macrolides in dairy goats. Small Rumin Res 167:55-60. DOI: https://doi.org/10.1016/j.smallrumres.2018.08.008

Robinson RK, Tamime AY, Wszolek M, 2002. Dairy microbiology handbook: the microbiology of milk and milk products, 3rd ed. ed. Wiley-Interscience, New York, NY, USA. DOI: https://doi.org/10.1002/0471723959

San Martin B, Kruze J, Morales MA, Agüero H, Leon B, Esppinoza S, Iragüen D, Puga J, Borie C, 2002. Resistencia bacteriana en cepas patógenas aisladas de mastitis en vacas lecheras de la V Región, Región Metropolitana y Xa Región, Chile. Arch Med Vet 34:2. DOI: https://doi.org/10.4067/S0301-732X2002000200008

Serhan M, Mattar J, Debs L, 2016. Concentrated yogurt (Labneh) made of a mixture of goats’ and cows’ milk: Physicochemical, microbiological and sensory analysis. Small Rumin Res 138:46-52. DOI: https://doi.org/10.1016/j.smallrumres.2016.04.003

Srinivasan V, Gillespie BE, Lewis MJ, Nguyen LT, Headrick SI, Schukken YH, Oliver SP, 2007. Phenotypic and genotypic antimicrobial resistance patterns of Escherichia coli isolated from dairy cows with mastitis. Vet Microbiol 124:319-28. DOI: https://doi.org/10.1016/j.vetmic.2007.04.040

Stolker AAM, Rutgers P, Oosterink E, Lasaroms JJP, Peters RJB, Van Rhijn JA, Nielen MWF, 2008. Comprehensive screening and quantification of veterinary drugs in milk using UPLC–ToF-MS. Anal Bioanal Chem 391:2309-22. DOI: https://doi.org/10.1007/s00216-008-2168-8

USAID, 2017. Processed foods value chain assessment report – Lebanon. USAID, Washington DC, USA.

Zanella GN, Mikcha JMG, Bando E, Siqueira VLD, Machinski M, 2010. Occurrence and antibiotic resistance of coliform bacteria and antimicrobial residues in pasteurized cow’s milk from Brazil. J Food Prot 73:1684-7. DOI: https://doi.org/10.4315/0362-028X-73.9.1684

Zeina K, Pamela AK, Fawwak S, 2013. Quantification of antibiotic residues and determination of antimicrobial resistance profiles of microorganisms isolated from bovine milk in Lebanon. Food Nutrition Sci 04:1-9. DOI: https://doi.org/10.4236/fns.2013.47A001